Bottom Line:
Activation of CCAP or CAMB neurons through temperature-sensitive TRPM8 gating is sufficient to trigger ecdysis behavior.Our findings demonstrate that kinin and CAMB neurons are direct targets of ETH and play critical roles in scheduling successive behavioral steps in the ecdysis FAP.Moreover, temporal organization of the FAP is likely a function of ETH receptor density in target neurons.

Mentions:
During the pupal ecdysis FAP in Drosophila, pre-ecdysis behavior (air bubble translocation, alternating, anteriorly-directed rolling contractions along the lateral edges of the abdomen) proceeds for ~10 min, whereupon a switch to ecdysis behavior (lateral swinging movements along with anteriorly directed peristaltic contractions) occurs. This leads within ~1–2 min to head eversion, after which rhythmic “ecdysis swinging” contractions continue for ~10 min [2,8]. In our previous study, we assessed functional consequences of ablating three central peptidergic ETHR ensembles defined by peptides they release (FMRFa, eclosion hormone, and CCAP) on the ecdysis FAP. Loss of FMRFa neurons had no effect on scheduling of the FAP, whereas loss of eclosion hormone neurons had a minor effect, producing a ~6 min delay in the switch from pre-ecdysis to ecdysis behavior. Loss of the entire CCAP neuron ensemble caused severe disruption of the FAP by abolishing the switch to ecdysis. Here we expand the dataset to include examination of additional ETHR ensembles in scheduling of the FAP using the Gal4-UAS system to drive expression of apoptosis genes (rpr, hid) for cell killing (CK) and assessed behavioral outcomes (Fig 1 and S1 Fig). Our findings confirm previous results and provide new evidence for disruption of the FAP through ablation of kinin, MIP, Burs, and Pburs ensembles. Kinin neurons are necessary for pre-ecdysis scheduling (see next section), while ablation of several CCAP neuron subsets abolishes the switch to ecdysis behavior. The smallest CCAP neuron subset is characterized by co-expression of myoinhibitory peptide (MIP), allatostatin-CC (Ast-CC), and bursicon, which we refer to as “CAMB” neurons (S2C Fig). Based on these results, we focused on kinin and CAMB ensembles for deeper analysis of the neural basis for pre-ecdysis and ecdysis scheduling.

Mentions:
During the pupal ecdysis FAP in Drosophila, pre-ecdysis behavior (air bubble translocation, alternating, anteriorly-directed rolling contractions along the lateral edges of the abdomen) proceeds for ~10 min, whereupon a switch to ecdysis behavior (lateral swinging movements along with anteriorly directed peristaltic contractions) occurs. This leads within ~1–2 min to head eversion, after which rhythmic “ecdysis swinging” contractions continue for ~10 min [2,8]. In our previous study, we assessed functional consequences of ablating three central peptidergic ETHR ensembles defined by peptides they release (FMRFa, eclosion hormone, and CCAP) on the ecdysis FAP. Loss of FMRFa neurons had no effect on scheduling of the FAP, whereas loss of eclosion hormone neurons had a minor effect, producing a ~6 min delay in the switch from pre-ecdysis to ecdysis behavior. Loss of the entire CCAP neuron ensemble caused severe disruption of the FAP by abolishing the switch to ecdysis. Here we expand the dataset to include examination of additional ETHR ensembles in scheduling of the FAP using the Gal4-UAS system to drive expression of apoptosis genes (rpr, hid) for cell killing (CK) and assessed behavioral outcomes (Fig 1 and S1 Fig). Our findings confirm previous results and provide new evidence for disruption of the FAP through ablation of kinin, MIP, Burs, and Pburs ensembles. Kinin neurons are necessary for pre-ecdysis scheduling (see next section), while ablation of several CCAP neuron subsets abolishes the switch to ecdysis behavior. The smallest CCAP neuron subset is characterized by co-expression of myoinhibitory peptide (MIP), allatostatin-CC (Ast-CC), and bursicon, which we refer to as “CAMB” neurons (S2C Fig). Based on these results, we focused on kinin and CAMB ensembles for deeper analysis of the neural basis for pre-ecdysis and ecdysis scheduling.

Bottom Line:
Activation of CCAP or CAMB neurons through temperature-sensitive TRPM8 gating is sufficient to trigger ecdysis behavior.Our findings demonstrate that kinin and CAMB neurons are direct targets of ETH and play critical roles in scheduling successive behavioral steps in the ecdysis FAP.Moreover, temporal organization of the FAP is likely a function of ETH receptor density in target neurons.